Genome Analysis Toolkit

Variant Discovery in High-Throughput Sequencing Data

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Developed in the Data Sciences Platform at the Broad Institute, the toolkit offers a wide variety of tools with a primary focus on variant discovery
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Calculate genotype posterior probabilities given family and/or known population genotypes

Category
Variant Evaluation and Refinement

Overview

Calculate genotype posterior probabilities given family and/or known population genotypes

This tool calculates the posterior genotype probability for each sample genotype in a VCF of input variant calls,
based on the genotype likelihoods from the samples themselves and, optionally, from input VCFs describing allele
frequencies in related populations. The input variants must possess genotype likelihoods generated by
HaplotypeCaller, UnifiedGenotyper or another source that provides unbiased genotype likelihoods.

Statistical notes

The AF field is not used in the calculation as it does not provide a way to estimate the confidence
interval or uncertainty around the allele frequency, unlike AN which does provide this necessary information. This
uncertainty is modeled by a Dirichlet distribution: that is, the frequency is known up to a Dirichlet distribution
with parameters AC1+q,AC2+q,...,(AN-AC1-AC2-...)+q, where "q" is the global frequency prior (typically q << 1). The
genotype priors applied then follow a Dirichlet-Multinomial distribution, where 2 alleles per sample are drawn
independently. This assumption of independent draws follows from the assumption of Hardy-Weinberg equilibrium (HWE).
Thus, HWE is imposed on the likelihoods as a result of CalculateGenotypePosteriors.

Inputs

A VCF with genotype likelihoods, and optionally genotypes, AC/AN fields, or MLEAC/AN fields.

(Optional) A PED pedigree file containing the description of the relationships between individuals.

Optionally, a collection of VCFs can be provided for the purpose of informing allele frequency priors. Each of
these resource VCFs must satisfy at least one of the following requirement sets:

AC field and AN field

MLEAC field and AN field

Genotypes

Output

A new VCF with the following information:

Genotype posteriors added to the FORMAT fields ("PP")

Genotypes and GQ assigned according to these posteriors (note that the original genotype and GQ may change)

Per-site genotype priors added to the INFO field ("PG")

(Optional) Per-site, per-trio joint likelihoods (JL) and joint posteriors (JL) given as Phred-scaled probability
of all genotypes in the trio being correct based on the PLs for JL and the PPs for JP. These annotations are added to
the FORMAT fields.

Notes

By default, priors will be applied to each variant separately, provided each variant features data from at least
10 called samples (no-calls do not count). SNP sites in the input callset that have a SNP at the matching site in
the supporting VCF will have priors applied based on the AC from the supporting samples and the input callset
unless the --ignoreInputSamples flag is used. If a site is not called in the supporting VCF, priors will be
applied using the discovered AC from the input samples unless the --discoveredACpriorsOff flag is used.
For any non-SNP sites in the input callset, flat priors are applied.

Usage examples

Refine genotypes based on the discovered allele frequency in an input VCF containing many samples

Caveat

If applying family priors, only diploid family genotypes are supported

CalculateGenotypePosteriors specific arguments

This table summarizes the command-line arguments that are specific to this tool. For more details on each argument, see the list further down below the table or click on an argument name to jump directly to that entry in the list.

Indices to use for the read inputs. If specified, an index must be provided for every read input and in the same order as the read inputs. If this argument is not specified, the path to the index for each input will be inferred automatically.

Validation stringency for all SAM/BAM/CRAM/SRA files read by this program. The default stringency value SILENT can improve performance when processing a BAM file in which variable-length data (read, qualities, tags) do not otherwise need to be decoded.

Use AC rather than MLEAC
By default the tool looks for MLEAC first, and then falls back to AC if MLEAC is not found. When this
flag is set, the behavior is flipped and the tool looks first for the AC field and then fall back to MLEAC or
raw genotypes.

Do not use discovered allele count in the input callset for variants that do not appear in the external callset.
Calculate priors for missing external variants from sample data -- default behavior is to apply flat priors

One or more genomic intervals to exclude from processing
Use this argument to exclude certain parts of the genome from the analysis (like -L, but the opposite).
This argument can be specified multiple times. You can use samtools-style intervals either explicitly on the
command line (e.g. -XL 1 or -XL 1:100-200) or by loading in a file containing a list of intervals
(e.g. -XL myFile.intervals).

Global Dirichlet prior parameters for the allele frequency
The global prior of a variant site -- i.e. the expected allele frequency distribution knowing only that N alleles
exist, and having observed none of them. This is the "typical" 1/x trend, modeled here as not varying
across alleles. The calculation for this parameter is (Effective population size) * (steady state mutation rate)

Use external information only
When this flag is set, only the AC and AN calculated from external sources will be used, and the calculation
will not use the discovered allele frequency in the callset whose posteriors are being calculated. Useful for
callsets containing related individuals.

Amount of padding (in bp) to add to each interval you are excluding.
Use this to add padding to the intervals specified using -XL. For example, '-XL 1:100' with a
padding value of 20 would turn into '-XL 1:80-120'. This is typically used to add padding around targets when
analyzing exomes.

Interval merging rule for abutting intervals
By default, the program merges abutting intervals (i.e. intervals that are directly side-by-side but do not
actually overlap) into a single continuous interval. However you can change this behavior if you want them to be
treated as separate intervals instead.

The --interval-merging-rule argument is an enumerated type (IntervalMergingRule), which can have one of the following values:

Amount of padding (in bp) to add to each interval you are including.
Use this to add padding to the intervals specified using -L. For example, '-L 1:100' with a
padding value of 20 would turn into '-L 1:80-120'. This is typically used to add padding around targets when
analyzing exomes.

Set merging approach to use for combining interval inputs
By default, the program will take the UNION of all intervals specified using -L and/or -XL. However, you can
change this setting for -L, for example if you want to take the INTERSECTION of the sets instead. E.g. to
perform the analysis only on chromosome 1 exomes, you could specify -L exomes.intervals -L 1 --interval-set-rule
INTERSECTION. However, it is not possible to modify the merging approach for intervals passed using -XL (they will
always be merged using UNION).
Note that if you specify both -L and -XL, the -XL interval set will be subtracted from the -L interval set.

The --interval-set-rule argument is an enumerated type (IntervalSetRule), which can have one of the following values:

UNION

Take the union of all intervals

INTERSECTION

Take the intersection of intervals (the subset that overlaps all intervals specified)

Number of hom-ref genotypes to infer at sites not present in a panel
When a variant is not seen in a panel, this argument controls whether to infer (and with what effective strength)
that only reference alleles were observed at that site. E.g. "If not seen in 1000Genomes, treat it as AC=0,
AN=2000". This is applied across all external panels, so if numRefIsMissing = 10, and the variant is absent in
two panels, this confers evidence of AC=0,AN=20.

Pedigree file for samples
See https://software.broadinstitute.org/gatk/documentation/article.php?id=7696 for more details on the PED
format. Note that each -ped argument can be tagged with NO_FAMILY_ID, NO_PARENTS, NO_SEX, NO_PHENOTYPE to
tell the GATK PED parser that the corresponding fields are missing from the ped file.

Indices to use for the read inputs. If specified, an index must be provided for every read input and in the same order as the read inputs. If this argument is not specified, the path to the index for each input will be inferred automatically.

Validation stringency for all SAM/BAM/CRAM/SRA files read by this program. The default stringency value SILENT can improve performance when processing a BAM file in which variable-length data (read, qualities, tags) do not otherwise need to be decoded.

The --read-validation-stringency argument is an enumerated type (ValidationStringency), which can have one of the following values:

Other callsets to use in generating genotype posteriors
Supporting external panels. Allele counts from these panels (taken from AC,AN or MLEAC,AN or raw genotypes) will
be used to inform the frequency distribution underlying the genotype priors. These files must be VCF 4.2 spec or later.